Table 1.
Binding Affinities of CXCL12-AF647 at NLuc/CXCR4 or NLuc/ACKR3 Measured by NanoBRET
| Cell Line or Membrane Preparation | pKd | n | Tagged Receptor Expression Level (fmol/μg membrane)b |
|---|---|---|---|
| TG NLuc/CXCR4 | (7.15 ± 0.04)a | – | – |
| TG NLuc/CXCR4 membranes | 7.55 ± 0.06 (7.61 ± 0.10)a | 3 | 90.67 ± 17.06 |
| CRISPR NLuc/CXCR4 HEK | 7.50 ± 0.04 | 3 | 1.45 ± 0.43 |
| CRISPR NLuc/CXCR4 HeLa | 7.58 ± 0.04 | 4 | 2.12 ± 0.95 |
| CRISPR HiBiT/CXCR4 | 7.49 ± 0.05 | 4 | ND |
| TG NLuc/ACKR3 HEK*** | 8.12 ± 0.10 | 5 | – |
| TG NLuc/ACKR3 membranes | 8.83 ± 0.06 | 4 | 52.9 ± 17.4 |
| CRISPR NLuc/ACKR3 HeLa | 8.77 ± 0.11 | 6 | 0.27 ± 0.14 |
TG cell lines indicate cells expressing transgenic exogenously expressed receptors, CRISPR cells lines indicate those expressing a genome-edited receptor under endogenous promotion. TG NLuc/CXCR4 or TG NLuc/ACKR3 membranes made from the respective cell lines. ***p < 0.001 was determined by a one-way ANOVA and indicate a significant difference between assay configurations in the binding affinity of CXCL12-AF647 to NLuc/ACKR3.
a
pKd values in parentheses from Caspar et al. (2018).
b
Values indicate mean ± SEM of five independent experiments performed in triplicate.